Submucosal glands in the cartilaginous airways are thought to play important roles in protecting the human lung from infection by secreting antibacterial factors and controlling the composition and viscosity of fluid in the airways. In diseases such as cystic fibrosis (CF), asthma, and chronic bronchitis, Submucosal glands expand in mass (hypertrophy) and/or abundance (hyperplasia), leading to excessive and abnormal mucus production. Genetic defects in serous cells of submucosal glands have also been hypothesized to be a contributing factor in lung disease in CF. In addition to playing a role in innate immunity of the airways, submucosal gland ducts have also recently been recognized as a potential protective niche for proximal airway stem cells. During the previous two funding cycles of this grant, our laboratory has focused on elucidating the mechanisms of submucosal gland development from proximal airway progenitors using human, ferret, and mouse model systems. The goal of these studies has been to better characterize the phenotype of glandular progenitor cells by understanding the transcriptional pathways that control the growth of glands in the airway during normal development. Since submucosal gland progenitors also appear to have the capacity to contribute to surface airway epithelial cell renewal, the biology of stem cells in both submucosal gland and surface epithelial compartments may be closely linked. We have previously demonstrated that a transcription factor, lymphoid enhancing factor-1 (Lef-1), is induced during the initial stages of airway progenitor/stem cell commitment to form submucosal glands and is absolutely required for glands to develop in the airway. More recently, we have begun to dissect the regulatory pathways that control transcriptional activation of the Lef-1 promoter in cell line models and glandular progenitors in transgenic mice. We have found that Wnt/(3-catenin pathways appear to play a critical role in regulating the Lef-1 promoter through a set of related HMG-box transcription factors (TCP and Sox). This proposal will use in vitro polarized airway models, ex vivo xenograft airway models, and genetic mouse models to study how Wnts, Noggin, and BMPs coordinate TCF/Sox regulation of the Lef-1 promoter during submucosal gland morphogenesis. Based on the inhibitory and enhancer functions of an identified Wnt-responsive element in the Lef-1 promoter, we hypothesize that Lef-1 regulation in glandular progenitors is controlled by both inhibitory (Sox and BMPs) and inductive (Wnt, TCF, and Noggin) signals. These hypotheses will be tested using viral vector systems and knockout mouse models capable of modulating the abundance of both extrinsic secreted factors and intrinsic transcription factors important for maintaining progenitor/stem cell phenotype and/or commitment to form glands. Lastly, this proposal will also attempt to evaluate whether TCF/Lef-1 pathways play a role in maintaining and/or mobilizing glandular duct stem cell niches for surface airway epithelial renewal following injury. Ultimately, this project will increase our understanding of stem cell phenotypes in the airway that have multipotent capacity for submucosal gland development and potentially also surface airway epithelial renewal. An increased understanding of submucosal gland morphogenesis may aid in the development of new therapeutic approaches to treat submucosal gland hyperplasia and hypertrophy in hypersecretory lung diseases. Furthermore, a greater knowledge of stem cell biology in the airway will greatly benefit the development genetic-based therapies for inhibited disease such as CF.